Technology-assisted and sutureless microvascular anastomoses: evidence for current techniques.

BACKGROUND: Since the birth of reconstructive microvascular surgery, attempts have been made to shorten the operative time while maintaining patency and efficacy. Several devices have been developed to aid microsurgical anastomoses. This article investigates each of the currently available technologies and attempts to provide objective evidence supporting their use. METHODS: Techniques of microvascular anastomosis were investigated by performing searches of the online databases Medline and Pubmed. Returned results were assessed according to the criteria for ranking medical evidence advocated by the Oxford Centre for Evidence Based Medicine. Emphasis was placed on publications with quantifiable endpoints such as unplanned return to theatre, flap salvage, and complication rates. RESULTS: There is a relative paucity of high-level evidence supporting any form of assisted microvascular anastomosis. Specifically, there are no randomized prospective trials comparing outcomes using one method versus any other. However, comparative retrospective cohort studies do exist and have demonstrated convincing advantages of certain techniques. In particular, the Unilink™/3M™ coupler and the Autosuture™ Vessel Closure System® (VCS®) clip applicator have been shown to have level 2b evidence supporting their use, meaning that the body of evidence achieves a level of comparative cohort studies. CONCLUSION: Of the available forms of assisted microvascular anastomoses, there is level 2b evidence suggesting a positive outcome with the use of the Unilink™/3M™ coupler and the Autosuture™ VCS® clip applicator. Other techniques such as cyanoacrylates, fibrin glues, the Medtronic™ U-Clip®, and laser bonding have low levels of evidence supporting their use. Further research is required to establish any role for these techniques.

Immunological responses after immunisation of mice with microparticles containing antigen and single stranded RNA (polyuridylic acid).

Certain toll-like receptor (TLR) agonists, e.g. CpG DNA, can be used as potent vaccine 'adjuvants'. It is known that some sequences of single stranded (ss) RNA stimulate proinflammatory and antiviral responses following interaction with TLR 7 and 8. We have encapsulated ovalbumin (OVA) in the presence and absence of polyuridylic acid (poly-U) inside polylactide microparticles. In comparison to microparticles containing only OVA, bulk cultures of bone marrow-derived plasmacytoid and myeloid dendritic cells produced more (P<0.05) IL-12 and interferon (IFN)-alpha when stimulated with microparticles containing OVA and poly-U. Subcutaneous injection of comicroencapsulated OVA and poly-U resulted in statistically elevated levels of serum anti-OVA IgG1 (P<0.05 versus naïve mice). Conversely, anti-OVA IgG1 levels in C57 BL6 mice immunised with OVA loaded microparticles (without RNA) were statistically indifferent to naïve animals. Furthermore, injection of coencapsulated OVA and poly-U resulted in (P<0.05) greater numbers of OVA specific IFN-gamma secreting T-cells as compared with mice injected with OVA loaded microparticles. A similar trend was seen in mice immunised with OVA loaded microparticles decorated with CpG or solutions of admixed OVA and CpG (P<0.05). These data demonstrate, for the first time, that appropriately formulated ssRNA can act as a potent adjuvant and modulator of adaptive immunological responses.

Protection against heterologous Burkholderia pseudomallei strains by dendritic cell immunization.

Burkholderia pseudomallei, the causative agent of melioidosis, is a gram-negative bacterium which can cause either chronic infections or acute lethal sepsis in infected individuals. The disease is endemic in Southeast Asia and northern Australia, but little is known about the mechanisms of protective immunity to the bacterium. In this study, we have developed a procedure to utilize dendritic cells in combination with CpG oligodeoxynucleotides as a vaccine delivery vector to induce protective immune responses to various strains of B. pseudomallei. Our results show that strong cell-mediated immune responses were generated, while antibody responses, although low, were detectable. Upon virulent challenge with B. pseudomallei strain K96243, NCTC 4845, or 576, animals immunized with dendritic cells that were pulsed with heat-killed K96243 and matured in the presence of CpG 1826 showed significant levels of protection. These results show that a vaccine strategy that actively targets dendritic cells can evoke protective immune responses.

Activation of dendritic cells by microparticles containing Bacillus anthracis protective antigen.

We have carried out an in vitro investigation into the mechanism by which microencapsulation enhances the immunogenicity of recombinant protective antigen (rPA) from Bacillus anthracis. Murine bone marrow derived dendritic cells (DC) were cocultured with soluble and microencapsulated rPA and the activation status of the cells monitored using FACS. As compared with soluble rPA, it was found that coculture of DC with rPA-loaded microparticles stimulated higher levels of MHC II, CD54, CD80 and CD86 expression (p<0.05). To investigate the longevity of antigen presentation, splenocytes from naïve mice were pulsed overnight with (3)H-thymidine following 1, 3 or 6 days coculture with DC transiently exposed to soluble or microencapsulated rPA. Splenocyte proliferation was more pronounced, and continued for a more protracted period, if the 'feeder' cells were exposed to microencapsulated antigen as compared with soluble antigen or 'empty' microspheres. To this end, our findings indicate that microsphere uptake increases the surface expression of MHC and co-stimulatory molecules on DC and can facilitate prolonged presentation of antigen to T-cells, possibly by acting as an intracellular depot.